Our microdialysis results showed that all tested doses of AMPH led to DA release in the CPu in a dose-dependent manner. The increased DA levels correlated to increased cAMP activity in the CPu and NAc (R2
> 0.92). On the other hand, the rCBV responses to the same four intravenous AMPH doses showed different patterns than the cAMP activity did. Although the rCBV responses in general agreed with the cAMP activity that the higher AMPH dose led to a stronger neuronal activity, there was a decoupling between the cAMP activity and rCBV response to the lower dose of AMPH (0.25mg/kg). Since only the postsynaptic DA receptors are linked to G-protein and cAMP activity, our cAMP data suggests that D1R has a stronger influence in the postsynaptic signal transduction than D2R does at doses of AMPH above 1mg/kg (or at approximately a 1000% increase in DA). Although the apparent affinity for dopamine appears to be similar for D1 and post-synaptic D2 receptor subtypes (Sokoloff et al., 1992
), the cAMP data indicates that, functionally, D1R has greater influence on the G-protein or cAMP activity than D2R does and thus linked to a positive rCBV contribution. The proportion of D1R influence increases as the DA concentration increases.
The D1R family includes D1 and D5 subtypes while the D2R receptor family includes D2, D3, and D4 subtypes. Among all of the DA receptor subtypes, the D2 autoreceptor and the D3R have highest affinity for dopamine and the D5R has higher affinity than the D1R (ki = 2300/2000/30nM for D1R/D2R/D3R) (Sokoloff et al., 1992
). Thus at lower DA concentrations, we can expect higher occupancy of the D2 autoreceptors and the D3R following stimulant drug administration (Richtand et al., 2001
). The presynaptic receptors (DA autoreceptor and DA transporter protein) do not couple to cAMP activity. Although there is some controversy on this topic, the D3R can act as a synthesis-modulating DA autoreceptor on the DA terminals (Aretha et al., 1995
). DA thus acts as a D3 agonist to suppress DA release from the presynaptic neurons. It is possible that, at low concentration (such as induced by 0.25mg/kg AMPH), DA primarily binds to D3R and the D2 autoreceptor more than to the postsynaptic DA receptors. The presynaptic neuronal activity thus dominates the majority of the gross neuronal activity and leads to rCBV decreases. When the DA level increases (by higher dose of AMPH), the proportion of synaptic DA binding to the postsynaptic receptors increases and the postsynaptic neuronal contribution overcomes the presynaptic one and thus rCBV increases.
One interesting phenomenon is that all AMPH doses led to rCBV increases in the lateral frontal cortex, despite the sign of rCBV in the CPu. The lateral frontal cortex is considered as one of the downstream structures in the DA circuitry and receives relatively little DA innervation directly. It is plausible that the rCBV increases in the lateral frontal cortex is the consequence of a positive “postsynaptic” neuronal activity in the major DA-innervating areas, even in the case of 0.25mg/kg AMPH that had positive “postsynaptic” cAMP activity but negative overall (presumably “presynaptic”) rCBV responses. In examining the maps produced by 0.25 mg/kg AMPH, it can be seen that in the anteromedial CPu there is a decreased CBV whereas in the posterior-lateral “motor” striatum there is an increased CBV. This would accord with the greater stimulated DA overflow seen in the latter region compared to the former region (Garris et al., 1994a
; Patel et al., 1992
). The increased DA overflow in the latter region would thus lead to an increased CBV compared to the anteromedial CPu as we observed. These observations suggest that the posterior-lateral striatal connections to the fronto-parietal cortex are more important in driving the CBV response, consistent with the mapping of the sensory-motor cortex to this region of the CPu (Heimer et al., 1995
One of the caveats is the direct coupling between the DA and vesculature. In addition to the receptor binding on the neurons, significant amount of DA is known to diffuse beyond synapses (Capella et al., 1993
; Garris et al., 1994b
) to regulate cerebrovascular smooth muscle (Edvinsson et al., 1985
). It has been shown that microapplication of DA in the vicinity of larger cerebral microvessels produced vasoconstriction (Edvinsson et al., 1985
; Krimer et al., 1998
). However, microapplication of D1 agonist led to vesodilation (Edvinsson et al., 1985
). Reverse transcription polymerase chain reaction (RT-PCR) studies for DA receptor subtypes on isolated cerebral microvessels with a size range from capillaries to aterioles with a diameter < 150µ showed only expression of D1R/D5R but not D2R/D3R (Choi et al., 2006
). In addition, D3R were found abundant in endothelial cells and astrocytes but not on microvessels (Choi et al., 2006
). It is likely that DA leads to vasodilation by binding to D1R on the microvessels and DA leads to vasoconstriction by binding to D3R on the glia cells to suppress the release of vasoactive molecules from astrocytes (Mulligan and MacVicar, 2004
; Simard et al., 2003
; Zonta et al., 2003
). Again, the negative rCBV response in the CPu in response to the lower dose of AMPH (0.25mg/kg) may due to a vasoconstriction effect via D3R binding since the DA binding affinity is much higher for D3R than the other DA receptor subtypes. However, currently we are not able to separate the hemodynamic effect driven by neuronal activity or a direct neurotransmitter-vasomotor coupling.
To summarize our results, we found that all doses tested of AMPH led to DA increases in the CPu. The DA concentrations were correlated to increased cAMP activity in the CPu and NAc, in a dose-dependent manner. AMPH also led to dose-dependent rCBV changes - rCBV increases from negative to positive as the dose of AMPH increases. We conclude that at low DA levels, the combined neuronal activity is dominated by the presynaptic activity. As the DA level increases, the postsynaptic activity gradually dominates the overall neuronal activity.